Genome-wide microRNA screening reveals miR-582-5p as a mesenchymal stem cell-specific microRNA in subchondral bone of the human knee joint

Abstract

Emerging evidence suggests that microRNAs (miRNAs) may be pathologically involved in osteoarthritis (OA). Subchondral bone (SCB) sclerosis is accounted for the knee osteoarthritis (KOA) development and progression. In this study, we aimed to screen the miRNA biomarkers of KOA and investigated whether these miRNAs regulate the differentiation potential of mesenchymal stem cells (MSCs) and thus contributing to SCB. We identified 48 miRNAs in the blood samples in KOA patients (n = 5) through microarray expression profiling detection. After validation with larger sample number, we confirmed hsa-miR-582-5p and hsa-miR-424-5p were associated with the pathology of SCB sclerosis. Target genes prediction and pathway analysis were implemented with online databases, indicating these two candidate miRNAs were closely related to the pathways of pluripotency of stem cells and pathology of OA. Surprisingly, mmu-miR-582-5p (homology of hsa-miR-582-5p) was downregulated in osteogenic differentiation and upregulated in adipogenic differentiation of mesenchymal progenitor C3H10T1/2 cells, whereas mmu-mir-322-5p (homology of hsa-miR-424-5p) showed no change through the in vitro study. Supplementing mmu-miR-582-5p mimics blocked osteogenic and induced adipogenic differentiation of C3H10T1/2 cells, whereas silencing of the endogenous mmu-miR-582-5p enhanced osteogenic and repressed adipogenic differentiation. Further mechanism studies showed that mmu-miR-582-5p was directly targeted to Runx2. Mutation of putative mmu-miR-582-5p binding sites in Runx2 3' untranslated region (3'UTR) could abolish the response of the 3'UTR-luciferase construct to mmu-miR-582-5p supplementation. Generally speaking, our data suggest that miR-582-5p is an important biomarker of KOA and is able to regulate osteogenic and adipogenic differentiation of MSCs via targeting Runx2. The study also suggests that miR-582-5p may play a crucial role in SCB sclerosis of human KOA.

Keywords: knee osteoarthritis; mesenchymal stem cells; miR-582-5p; subchondral bone.